Information about genes is critical to understanding the biological processes which underlie life: cellular communication, growth, movement, reproduction, and control. Once obtained, the structural and functional features of the genetic sequences (polypeptides and polynucleotides) enable better diagnostics and treatment of disease and defects, whether genetic or external in origin.
Traditional gene expression research has involved manual pipetting of samples onto gels, membranes or filters, or into multiwell plates. These methodologies are extremely time consuming, laborious, low throughput and expensive (on a per gene basis).
Modern microarray technologies get around a number of the above problems by automating the spotting process using robotics which permits high density spotting of slides, also known as microarray slides, which allows thousands of gene fragments to be analyzed in a single experiment (Schena M, Shalon D, Davis R W, Brown PO. “Quantitative monitoring of gene expression patterns with a complementary DNA microarray”, Science 270, 467470 (1995); Southern, E, Mir K, and Shchepinov, M, Molecular Interactions on microarrays, Nature Genet. 21, 5-9 (1999)). Each spot leaves a sample of volume in the nanoliter range, the centres of adjacent spots separated by micrometers.
Array types include oligonucleotide arrays, cDNA arrays and genomic DNA arrays. For example, one application is to identify the genes, the expression or repression of which results in the difference between a normal human cell and a mutant human cell. Cells contain thousands of genes, a few thousand in lower organisms such as yeast and over 100,000 in humans; a cDNA is made for each gene and spotted onto gene chips as part of the microarray. Another application concerns the construction of olionucleotide arrays. It is also possible for genomic DNA arrays (chromosomal DNA) to be fabricated using modern microarray technology. Arrays are also useful for DNA sequencing.
Typically, a microarrayer has a number of components including: (1) a robotic mechanism; (2) a dispenser assembly; (3) means for replenishing the spotting dispensers with the biological sample; (4) a platform to support the microarray slides during spotting; (5) means for cleaning the spotters; and (6) software to operate the robot mechanism and provide an interface with a user. Samples are typically stored in source plates. Source plate plastics may be polystyrene, polypropylene or polycarbonate. In general, plates may have 1536, 384 or 96 wells. DNA may be attached to the substrate through any suitable technique (e.g. covalent or ionic bonding). Dispensers for spotting, also known as spotting members, include pins, which in turn include solid pins, split or quill pins, pin and ring systems, capillaries, or inkjet systems. Pins include the Telechem Chipmaker 2 and the Telechem Chipmaker 3 (Stealth Pins).
Recent advances in robotics such as that disclosed by the invention in U.S. Pat. No. 6,048,373, have made possible a number of features desired in an ideal microarrayer: (1) high resolution; (2) repeatability; and (3) precision. Resolution of a system is the ability to distinguish two points as being separate; it is also the minimum distance that can be measured by an encoder. Repeatability refers to the ability of the robot to return to the same place. The difference between the position that the robot desires to occupy and the actual position occupied is the precision of the system. The density of the array is a function of the spatial resolution of the robot.
The mechanism for dispensing the biological sample typically uses pins as the part of the print head that performs the actual spotting. The preferred approach is a set of pins, either in the solid or in the split form of the pin (with a slot), typically arranged in a rectangular matrix. The biological samples are loaded into/onto the pins from the source plates. It is critical that deposition of probe biological sample, such as cDNA, yield regularly spaced spots of uniform morphology. Not all deposition pins designed to the same specifications behave in a similar manner. Each will load an amount characteristic of the pin. Consequently, the size of the first spots produced from a set of pins can be significantly variable. The greatest concern is that deposition of excessive material on the microarrays may yield overlapping spots. This will result in contamination of the material spotted on the arrays as well as the material in the probe plates. As spotting proceeds, the excess material is removed and the spots become uniform until the exhaustion of the material on the pin. There is thus a need to effectively remove the excess material prior to spotting onto the microarray slides.
It is also important to have a well-designed platform (also known as a platen) onto which the microarray slides are placed for array printing. Existing art uses a platform that had “rails” cut into it. These rails would serve to hold the slides in place in the X-axis and/or Y-axis, and allow for just enough space into which to place the slides; however in certain cases, the lack of a uniform standard on slide sizes means that certain commercially available slides would not fit into the tracks of a particular microarrayer. The rails have to be machined with great precision to hold the slides without allowing for movement. Other solutions use a set of spring loaded plastic pins, which hold the slides in place. These pins offer some compliance to allow for subtle variations in slide size (such as the difference between metric and imperial measure slides). Other units utilize a combination of machined holders for the slides with a vacuum manifold. The vacuum manifold helps hold the slides down firmly which allows the depression into which the slide sits to not be an exact fit. Again in this case slight size variations are possible allowing imperial and metric slides to be used.
These solutions do not provide for a great deal of flexibility. In addition, these slide platforms/holders are difficult to load, which causes the operation to be time consuming and increases the risk of damage to the slides. Some of these units require such a platform/holder design because the slide platform sits on top of one of the robotic actuators. In such a case the slide platform moves during the printing process and thus the slides need to be held firmly to prevent them from shifting in place. Where the robot mechanism uses an overhead gantry system; the print head travels over the slide platform in all three axes, and the slide platform remains stationary. In such a situation, there is little movement, which will cause the slides to shift. There is thus a need for a platform/slide holder, which allows for much greater flexibility, and much greater ease of use.
In order to clean the spotting pins, a vacuum or forced air removes liquid from the pins, usually present on the pins (and in the slot of the pins) as a result of dipping into a water bath for cleaning after spotting. Known vacuum manifolds are comprised of a chamber containing a series of holes (as many as there are pins for the print head) into which the pins fit. The tips of pins are generally placed into the holes, either at the opening of the hole, or completely into the vacuum chamber. These solutions are not sufficient for optimal cleaning of the pins. There is a need for a vacuum manifold and associated method to optimally clean the spotting pins.